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Real-Time Imaging of Intracellular Glutathione Levels Based on a Ratiometric Fluorescent Probe with Extremely Fast Response.
Analytical Chemistry ( IF 6.7 ) Pub Date : 2020-06-15 , DOI: 10.1021/acs.analchem.0c01881
Ming Tian 1 , Xing-Yu Liu 1 , Huan He 1 , Xian-Zheng Ma 2 , Chao Liang 2 , Yi Liu 1, 3, 4 , Feng-Lei Jiang 1
Affiliation  

Glutathione (GSH), the most abundant nonprotein thiol found in living organisms, are involved in the etiology and progression of many human diseases including cancer. So, monitoring changes of cellular GSH levels has an important guiding significance. To date, however, majority of probes can only qualitatively detect GSH in living cells. Herein, with coumarin as the read-out fluorophore and Michael addition as the sensing mechanism, six fluorescent probes were designed and synthesized. Among them, RP-2 exhibited a reversible and extremely fast response toward GSH (half time: ∼3 s), which endowed RP-2 the capacity of real-time imaging. Among the reversible probes based on Michael addition, RP-2 had both the largest forward and reverse rate constants thus far. The reaction between RP-2 and GSH was studied in detail by density functional theory and fluorescence spectroscopy. Real-time imaging of GSH levels in living cells was achieved with a temporal resolution of seconds. To simplify the processing of images, a program was developed and validated. RP-2 was expected to serve as a new fluorescent imaging tool to understand the function of intracellular GSH in the future.

中文翻译:

基于具有极快响应的比例荧光探针对细胞内谷胱甘肽水平的实时成像。

谷胱甘肽(GSH)是在活生物体中发现的最丰富的非蛋白质硫醇,它参与许多人类疾病(包括癌症)的病因和病情发展。因此,监测细胞谷胱甘肽水平的变化具有重要的指导意义。然而,迄今为止,大多数探针只能定性检测活细胞中的谷胱甘肽。在此,以香豆素为读出的荧光团并以迈克尔加成法为感测机制,设计并合成了六个荧光探针。其中,RP-2对GSH表现出可逆且极快的响应(一半时间:约3 s),这赋予了RP-2实时成像的能力。在基于迈克尔加法的可逆探针中,RP-2具有迄今为止最大的正向和反向速率常数。通过密度泛函理论和荧光光谱研究了RP-2和GSH之间的反应。活细胞中GSH水平的实时成像可在数秒的时间分辨率下实现。为了简化图像处理,开发并验证了程序。RP-2有望作为一种新的荧光成像工具来了解未来细胞内GSH的功能。
更新日期:2020-07-21
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